Characterization of isolates of Ralstonia solanacearum biovar 2, pathogenic to Eucalyptus “urograndis” hybrids

ABSTRACTThe objective of this study was to characterize isolates of biovar 2 of Ralstonia solanacearum pathogenic to eucalyptus “urograndis” hybrid by means of biochemical tests, as well as evaluation of host range and identification by PCR with primers for species, biovar and phylotype. Carbohydrate utilization assays revealed that the eucalyptus isolates belong to phenotype Tropical of biovar 2 (2T). From artificial inoculations it was possible to reproduce symptoms or recover the bacterium from: eucalyptus, potato, tomato, eggplant, datura, geranium, turnip, mustard, nasturtium, beetroot, sunflower, bean, French marigold, horseradish tree, and cashew. The PCR assays confirmed that the isolates belong to biovar 2, phylotype II of the bacterium. The confirmation of new biovar naturally infecting eucalyptus indicates that R. solanacearum easily adapts to new hosts

Controle biológico do cancro da teca causado por Lasiodiplodia theobromae

A teca é uma espécie florestal que assumiu grande importância no Brasil, onde encontrou excelentes condições de desenvolvimento desde a sua introdução na década de 1960. Entretanto, problemas fitossanitários começam a ameaçar a exploração desta espécie madeireira. Um exemplo é o cancro da teca, causado pelo fungo Lasiodiplodia theobromae (Lt), cuja etiologia foi recentemente elucidada no Brasil e, portanto, ainda não existem métodos de controle recomendados. Sendo assim, o objetivo deste trabalho foi avaliar o controle deste patógeno, investigando o potencial dos agentes de controle biológico (ACB) Trichoderma spp., Bacillus sp. e Enterobacter sp., inicialmente por meio de ensaios in vitro e, posteriormente, com realização de testes in vivo. De acordo com os resultados dos testes in vitro, os isolados de Trichoderma CEN162 e CEN1153 e a estirpe UnB1366 de Bacillus sp., foram os tratamentos que se destacaram, sendo capazes de inibir completamente o crescimento micelial de alguns isolados de Lt. Quando esses isolados foram testados de forma preventiva, observou-se variação nos níveis de controle, dependendo do isolado de Lt e da interação antagonista-clone, onde CEN162 (T. asperellum) e UnB166 (Bacillus sp.) apresentaram 100% de controle. Dessa forma, houve uma correlação positiva entre os testes in vitro e in vivo, uma vez que os mesmos ACB se destacaram. Apesar de ter-se obtido bons níveis de controle com os ACB utilizados, pode-se concluir que existe uma variação no antagonismo a diferentes isolados ou mesmo na interação antagonista-clone, corroborando as informações disponíveis na literatura científica sobre este fungo fitopatogênico.Teak is a forest species that has assumed great importance in Brazil, where it has found excellent conditions for development since its introduction into the country in the 1960s. However, phytosanitary problems are beginning to threaten the production of this timber species. An example is teak canker, caused by the fungus Lasiodiplodia theobromae (Lt), which has only recently been reported in Brazil, and for which, therefore, there are no recommended control methods. Thus, this study evaluated the control of this pathogen, investigating the potential of the biocontrol agents (BCAs) Trichoderma spp., Bacillus sp. and Enterobacter sp., initially through in vitro assays and, subsequently, with in vivo tests. According to the in vitro assay results, the Trichoderma isolates CEN162 and CEN1153 and the strain of Bacillus sp. (UnB1366) were the treatments that stood out, as they were able to completely inhibit mycelial growth of some isolates of Lt. When these isolates were tested in a preventive way, the control levels varied depending on the Lt isolate and the antagonist-clone interaction, where CEN162 (T. asperellum) and UnB166 (Bacillus sp.) showed 100% control. Thus, there is a positive correlation between the in vitro and in vivo tests, since the same BCAs stood out. Although good levels of control have been obtained with the BCAs used, it can be concluded that there is a variation in the antagonism to different Lt isolates or even in the antagonist-clone interaction, corroborating the information available in the scientific literature on this plant-pathogenic fungus

Novos isolados de Trichoderma antagônicos a Sclerotinia sclerotiorum

Quarenta e nove isolados de Trichoderma obtidos no centro-oeste do Brasil foram avaliados quanto a sua atividade antagônica in vitro contra Sclerotinia sclerotiorum (agente causal do mofo branco) e identificados com base nas sequências ITS do DNA ribossômico nuclear. Os testes de cultivo pareado mostram que todos os isolados exibiram algum antagonismo, com um máximo de 77% de inibiação micelial e inibição total da produção de escleródios. Dois isolados se destacaram como os mais promissores, considerando ambos os parâmetros avaliados (CEN1253 - T. koningiopsis e CEN1265 - T. brevicompactum). Cinco espécies diferentes foram identificadas: T. harzianum (23), T. spirale (9), T. koningiopsis (8), T. brevicompactum (7) and T. asperellum (2). Estes isolados estão armazenados na Coleção de Fungos para Controle Biológico da Embrapa e as informações obtidas nos experimentos serão incorporadas na base de dados de ativos biológicos, no sistema de informações de recursos genéticos, e disponibilizados para estudos futuros.Forty-nine isolates of Trichoderma from the Brazilian Midwest were evaluated for their antagonistic activity in vitro against Sclerotinia sclerotiorum (causal agent of white mold), which were then identified based on their nuclear ribosomal ITS sequences. Paired culture tests showed that all isolates exhibited some antagonism, with a maximum of 77% mycelial inhibition and complete inhibition of sclerotia production. Two isolates were found to be the most promising biocontrol agents, considering both antagonistic parameters (CEN1253 - T. koningiopsis and CEN1265 - T. brevicompactum). Five different species were identified: T. harzianum (23), T. spirale (9), T. koningiopsis (8), T. brevicompactum (7) and T. asperellum (2). These isolates are stored in the Embrapa Fungi Collection for Biological Control and the information obtained in the experiments will be incorporated into the database of biological assets within the genetic resources information system (Allele) and be made available for further studies

Functional and evolutionary analyses of the miR156 and miR529 families in land plants

Abstract\ud \ud Background\ud MicroRNAs (miRNAs) are important regulatory elements of gene expression. Similarly to coding genes, miRNA genes follow a birth and death pattern of evolution likely reflecting functional relevance and divergence. For instance, miRNA529 is evolutionarily related to miRNA156 (a highly conserved miRNA in land plants), but it is lost in Arabidopsis thaliana. Interestingly, both miRNAs target sequences overlap in some members of the SQUAMOSA promoter-binding protein like (SPL) family, raising important questions regarding the diversification of the miR156/miR529-associated regulatory network in land plants.\ud \ud \ud Results\ud In this study, through phylogenic reconstruction of miR156/529 target sequences from several taxonomic groups, we have found that specific eudicot SPLs, despite miRNA529 loss, retained the corresponding target site. Detailed molecular evolutionary analyses of miR156/miR529-target sequence showed that loss of miR529 in core eudicots, such as Arabidopsis, is correlated with a more relaxed selection of the miRNA529 specific target element, while miRNA156-specific target sequence is under stronger selection, indicating that these two target sites might be under distinct evolutionary constraints. Importantly, over-expression in Arabidopsis of MIR529 precursor from a monocot, but not from a basal eudicot, demonstrates specific miR529 regulation of AtSPL9 and AtSPL15 genes, which contain conserved responsive elements for both miR156 and miR529.\ud \ud \ud Conclusions\ud Our results suggest loss of functionality of MIR529 genes in the evolutionary history of eudicots and show that the miR529-responsive element present in some eudicot SPLs is still functional. Our data support the notion that particular miRNA156 family members might have compensated for the loss of miR529 regulation in eudicot species, which concomitantly may have favored diversification of eudicot SPLs.We thank Dr. Scott Poethig for 35S::AtMIR156a seeds; Dr. Peter Huijser for\ud spl9;spl15 seeds; Dr. Renato Vicentini for initial bioinformatic analyses and\ud helpful discussions; and Dr. Luiz Del Bem for initial phylogenetic analyses.\ud This work was supported by the State of Sao Paulo Research Foundation,\ud FAPESP, Brazil (grants no. 07/58289-5 and 12/51146-2). EGOM was a recipient\ud of a fellowship from Coordination for the Improvement of Higher Education\ud Personnel (CAPES, Brazil). GFFS (from Centro de Energia Nuclear na\ud Agricultura –CENA/USP) and EMS were recipients of a fellowship from the\ud State of Sao Paulo Research Foundation, FAPESP, Brazil